The rockefeller university

Over 500 primary immunodeficiency diseases (PID) have been described, but immunological assessment after a severe infection is not routine. We aimed to evaluate the feasibility of a PID screening protocol and calculate PID prevalence in children admitted for severe infection in a pediatric intensive care unit (PICU). This monocentric retrospective study evaluated the feasibility of a PID monitoring protocol after severe infection in children aged 1 month to 16 years-old hospitalized in the Montpellier University Hospital from January 2018 to December 2020. Follow-up consultations at 3 and 12 months included the three main PID screening scores, comprehensive immunological and genetic screenings. Among 1125 children admitted to the PICU, 46 had severe infections and caused by bacterial (48%), viral (39%) or fungal (2%) pathogens. Before infection, none had completed any screening score recommended by dedicated societies (Jeffrey Modell Foundation, German Patients' Organization for Primary Immunodeficiencies, French Reference Center for Hereditary Immunodeficiencies). At 3 months, three patients had a PID diagnosis (6.5% prevalence, 95% CI 1.4-17.9). These were associated with a deletion of chromosomal region 22q11.21 (DiGeorge syndrome), ELANE mutation (Elastase deficiency or Severe Congenital Neutropenia 1), and C5 deficiency Forty children (87%) presented immunological anomalies without a formal PID diagnosis. These persisted in only 4/17 children tested at 12 months. The most frequent abnormalities were low NK lymphocytes (41.18%), and abnormal B lymphocyte population distribution (25%). The observed PID prevalence post-severe infection matches previous reports, even with a high rate of viral infections, often overlooked. Systematic PID investigation after severe infection, regardless of the pathogen, should be implemented to improve early detection and treatment.

To produce a murine monoclonal antibody (mAb) that binds to glycoprotein IIb/IIIa (alpha IIb beta 3) and inhibits clot retraction (CR), we immunized mice with human platelets and tested hybridoma supernatants for their ability to bind to alpha IIb beta 3 and inhibit CR. The immunoglobulin G1 (IgG1) mAb R6H8 completely inhibited CR at 20 mu g/mL. Paradoxically, at 5 mu g/mL, R6H8 initiated platelet aggregation and induced P-selectin expression, fibrinogen binding, and PAC-1 binding. At 20 mu g/mL, however, R6H8 completely inhibited aggregation induced by thrombin PAR-1 receptor activating peptide SFLLRN (T6; 25 mu g/mL) and T6-induced fibrinogen and PAC-1 binding to platelets. Platelet aggregation induced by R6H8 was inhibited by mAb IV.3, which blocks the Fc gamma IIa receptor (Fc gamma RIIa), and the Fab fragment of R6H8 did not induce platelet aggregation, suggesting that R6H8 binds to both alpha IIb beta 3 and Fc gamma RIIa. Cryogenic electron microscopy analysis of the R6H8 Fab-alpha IIb beta 3 complex revealed that R6H8 (1) binds to the alpha IIb beta 3 RGD binding pocket via an Arg-Tyr-Asp (RYD) sequence in its heavy chain complementarity-determining region 3; (2) interacts with beta 3 Asp126, producing a reorientation of Asp126 and loss of the adjacent to metal ion-dependent adhesion site Ca2+; and (3) initiates swing-out of the beta 3 hybrid domain. We conclude that R6H8 is an alpha IIb beta 3 ligand-mimetic mAb that activates platelets via Fc gamma RIIa at low concentrations and potently inhibits platelet aggregation and CR at high concentrations. R6H8 simulates the actions of a number of pathological antibodies, including platelet-activating antibodies developed after therapy with alpha IIb beta 3 inhibitors and platelet-activating antibodies in heparin-induced thrombocytopenia and vaccine-induced immune thrombotic thrombocytopenia. As such, it may be a valuable reagent for better understanding these disorders and identifying potential therapies.

Proper fuelling of the brain is critical to sustain cognitive function, but the role of fatty acid (FA) combustion in this process has been elusive. Here we show that acute block of a neuron-specific triglyceride lipase, DDHD2 (a genetic driver of complex hereditary spastic paraplegia), or of the mitochondrial lipid transporter CPT1 leads to rapid onset of torpor in adult male mice. These data indicate that in vivo neurons are probably constantly fluxing FAs derived from lipid droplets (LDs) through beta-oxidation to support neuronal bioenergetics. We show that in dissociated neurons, electrical silencing or blocking of DDHD2 leads to accumulation of neuronal LDs, including at nerve terminals, and that FAs derived from axonal LDs enter mitochondria in an activity-dependent fashion to drive local mitochondrial ATP production. These data demonstrate that nerve terminals can make use of LDs during electrical activity to provide metabolic support and probably have a critical role in supporting neuron function in vivo.

Isocitrate dehydrogenase 1/2 (IDH) mutations are early initiating events in acute myeloid leukemia (AML). The complex clonal architecture and cellular heterogeneity in IDH-mutant AML underlies the heterogeneous clinical presentation and outcomes. Integrating single-cell genotyping and transcriptomics, we demonstrate a stem-like and inflammatory phenotype of IDH-mutant AML and identify clone-specific programs associated with NPM1, NRAS, and SRSF2 co-mutations. Furthermore, these clones had distinct responses to treatment with combination IDH inhibitors and chemotherapy, including elimination, reconstitution of myeloid differentiation, or retention within progenitor populations. At relapse after IDH inhibitor monotherapy, we identify up-regulated stemness, inflammation, mitochondrial metabolism, and anti-apoptotic factors, as well as down-regulated major histocompatibility complex (MHC) class II antigen presentation. At the pre-leukemic stage, we observe upregulation of IDH2-associated pathways, including inflammation. We deliver a detailed phenotyping of IDH-mutant AML and a framework for dissecting contributions of recurrently mutated genes in AML at diagnosis and following therapy, with implications for precision medicine.

Fanconi anemia (FA) is a rare genetic disease characterized by loss-of-function variants in any of the 22 previously identified genes (FANCA-FANCW) that encode proteins participating in the repair of DNA interstrand crosslinks (ICLs). Patient phenotypes are variable but may include developmental abnormalities, early-onset pancytopenia, and a predisposition to hematologic and solid tumors. Here, we describe 2 unrelated families with multiple pregnancy losses and offspring presenting with severe developmental and hematologic abnormalities leading to death in utero or in early life. Homozygous loss-of-function variants in FAAP100 were identified in affected children of both families. The FAAP100 protein associates with FANCB and FANCL, the E3 ubiquitin ligase responsible for the monoubiquitination of FANCD2 and FANCI, which is necessary for FA pathway function. Patient-derived cells exhibited phenotypes consistent with FA. Expression of the WT FAAP100 cDNA, but not the patient-derived variants, rescued the observed cellular phenotypes. This establishes FAAP100 deficiency as a cause of FA, with FAAP100 gaining an alias as FANCX. The extensive developmental malformations of individuals with FAAP100 loss-of-function variants are among the most severe across previously described FA phenotypes, indicating that the FA pathway is essential for human development.

Nanodiscs, small discoidal membrane patches stabilized by membrane-scaffold proteins (MSPs), are popular tools to stabilize integral membrane proteins (IMPs) for structural studies by cryogenic electron microscopy (cryo-EM). While nanodiscs provide a near-native membrane environment for the incorporated IMPs, they do not reproduce all characteristics of a native membrane. Also, IMPs must first be purified in detergent before they can be reconstituted into MSP-based nanodiscs, a problem that has been overcome by newer approaches, such as copolymer-based native nanodiscs and cell-derived vesicles. In this review, we argue that despite these advances, MSP-based nanodiscs remain a unique tool for the structural interrogation of IMPs.

Despite waning of virus-neutralizing antibodies, protection against severe SARS-CoV-2 in the majority of immune individuals remains high, but the underlying immune mechanisms are incompletely understood. Here, rhesus macaques with pre-existing immunity from Novavax WA-1 and/or P.1 vaccines and WA-1 or P.1 infection are immunized with a bivalent WA-1/Omicron BA.5 Novavax vaccine ten months after the last exposure. The boost vaccination primarily increases the frequency of cross-reactive spike (S)-specific antibodies and B cells instead of inducing de novo BA.5-specific responses. Reinfection with heterologous Omicron XBB.1.5 six months after the boost vaccination results in low levels of virus replication in the respiratory tract compared with virus-na & iuml;ve results from other studies. Whereas systemic S-specific immunity remains largely unchanged in all animals, the animals with complete protection from infection exhibit a stronger influx of S-specific IgG, monocytes, B cells and T cells into the bronchioalveolar space combined with expansion of CD69+CD103+ lung tissue-resident, S-specific CD8 T cells compared to actively infected animals. Our results underscore the importance of localized respiratory immune responses in mediating protection from Omicron reinfection and provide guidance for future vaccine development.

Defining viral proteomes is crucial to understanding viral life cycles and immune recognition but the landscape of translated regions remains unknown for most viruses. We have developed massively parallel ribosome profiling (MPRP) to determine open reading frames (ORFs) across tens of thousands of designed oligonucleotides. MPRP identified 4208 unannotated ORFs in 679 human-associated viral genomes. We found viral peptides originating from detected noncanonical ORFs presented on class-I human leukocyte antigen in infected cells and hundreds of upstream ORFs that likely modulate translation initiation of viral proteins. The discovery of viral ORFs across a wide range of viral families-including highly pathogenic viruses-expands the repertoire of vaccine targets and reveals potential cis-regulatory sequences.

Tauopathies are characterized by the aggregation and accumulation of hyperphosphorylated tau proteins that correlates with cognitive impairment in affected individuals. The presence of tauopathy follows a temporospatial spreading pattern in which certain neuronal cell types in specific brain regions are more vulnerable to tau accumulation and atrophy. However, the mechanisms underlying the selective vulnerability of these neurons and regions to pathological tau accumulation are not fully understood. Here, we characterized the presence of phosphorylated tau in excitatory and inhibitory neurons in post-mortem prefrontal cortex of tauopathy patients, including Alzheimer's disease, progressive supranuclear palsy, corticobasal degeneration, and frontotemporal lobar dementia due to a MAPT mutation. We observed that neuronal tau accumulation across these tauopathies occurs predominantly in excitatory neurons compared to inhibitory neurons. Next, we performed viral translating ribosome affinity purification (vTRAP) from vulnerable and resistant brain regions on vGLUT1CRE+ and GAD2CRE+ PS19 mice to understand molecular signatures of tau vulnerability. We observed that both vulnerable regions and vulnerable neurons are characterized by alterations in synaptic transmission and neuronal excitability. Transcription factor Mef2c (myocyte enhancer factor 2c) was identified as an upstream regulator affecting myelination and synaptic organization in vulnerable brain regions in PS19 mice. The relevance of these findings was validated in human tauopathies via coexpression network analysis. Concordantly, we observed tau-induced changes in spontaneous postsynaptic currents of excitatory neurons in mice especially in the prefrontal cortex. Taken together, we conclude that selective vulnerability to tau could arise from changes in neurotransmission and synaptic compositions, potentially due to an altered Mef2c transcriptional network.

The mammalian cochlea benefits from an active process characterized by amplification of mechanical inputs, sharp frequency selectivity, compressive nonlinearity, and spontaneous otoacoustic emission. Similar traits are observed in individual hair cells of nonmammalian tetrapods, in which they emerge from the critical dynamical regime of hair cells operating near a Hopf bifurcation. It remains unclear whether a similar critical regime also underpins the active process of the mammalian cochlea. Efforts to address this question have been limited in part by the absence of an ex vivo preparation that both preserves the physiological integrity of the sensory epithelium and grants direct experimental access to it. To overcome these problems, we improved a two-compartment cochlear preparation (Chan and Hudspeth, 2005a, 2005b) to more closely simulate in vivo conditions and used it to conduct electrophysiological recordings of microphonic signals in isolated cochlear segments of the Mongolian gerbil. Our methodological advances included refining the dissection protocol to reduce the size of the exposed cochlear segment and altering the ionic compositions of the solutions to better control the Ca2+ concentration. We also maintained a constant temperature in order to stabilize the experimental conditions. Most critically, by introducing a mechanism to adjust the pressure in the endolymphatic compartment, we were able to explore how variations in transepithelial pressure influence the electrical response. These changes enabled us to reliably measure compressive nonlinearities with a one-third power law similar to that observed from cochleas in vivo and consistent with the behavior of a dynamical system operating near a Hopf bifurcation.